LOCUS       sequence-325955-        8564 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV-bpNLS-dCas13d-bpNLS-ADARdd, ADARdd embeded in loop1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8564)
  AUTHORS   Liu Y, Mao S, Huang S, Li Y, Chen Y, Di M, Huang X, Lv J, Wang X, Ge
            J, Shen S, Zhang X, Liu D, Huang X, Chi T
  TITLE     REPAIRx, a specific yet highly efficient programmable A > I RNA base
            editor.
  JOURNAL   EMBO J. 2020 Nov 16;39(22):e104748. doi: 10.15252/embj.2020104748. 
            Epub 2020 Oct 15.
  PUBMED    33058207
REFERENCE   2  (bases 1 to 8564)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "EMBO J."; 
            date: "2020-11-16"; volume: "39(22):e104748. doi"; pages: " 
            10.15252/embj.2020104748. Epub 2020 Oct 15"
COMMENT     Sequence Label: TC1464
FEATURES             Location/Qualifiers
     source          1..8564
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(57..645)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(137..156)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(816..1607,1608..1676))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1439..1458
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1677..1781)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1856..1875)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2047..2426
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2427..2630
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2581..2601
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2675..2694
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2675..2693
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2771..2791
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3389..3409
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4610..4630
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7022..7042
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7049..7114
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             7139..7195
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             7202..7912
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     7615..7634
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(7951..7968)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7957..8181
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(8219..8235)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8219..8235)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8232..8254)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8243..8259
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(8267..8273,8274..8291,8292..8297))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8312..8333
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8450..8467)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
       61 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
      121 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
      181 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
      241 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
      301 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
      361 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
      421 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
      481 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
      541 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
      601 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
      661 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
      721 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
      781 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
      841 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
      901 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
      961 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     1021 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     1081 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     1141 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     1201 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     1261 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     1321 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     1381 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     1441 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     1501 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     1561 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     1621 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     1681 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     1741 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     1801 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
     1861 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
     1921 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
     1981 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
     2041 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
     2101 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
     2161 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
     2221 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
     2281 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
     2341 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
     2401 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
     2461 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
     2521 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
     2581 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa
     2641 ctagagaacc cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc
     2701 aagctggcta gcgtttaaac ttaagcttgc caccatgaaa cggactgctg acggcagcga
     2761 atttgagagc cccaagaaga agaggaaagt cggatccatc gagaagaaga agagcttcgc
     2821 caagggcatg ggagtgaaga gcaccctggt gtccggctct aaggtgtaca tgaccacatt
     2881 tgctgaggga agcgacgcca ggctggagaa gatcgtggag ggcgatagca tcagatccgt
     2941 gaacgaggga gaggctttca gcgccgagat ggctgacaag aacgctggct acaagatcgg
     3001 aaacgccaag ttttcccacc caaagggcta cgccgtggtg gctaacaacc cactgtacac
     3061 cggaccagtg cagcaggaca tgctgggact gaaggagaca ctggagaaga ggtacttcgg
     3121 cgagtccgcc gacggaaacg ataacatctg catccaggtc atccacaaca tcctggatat
     3181 cgagaagatc ctggctgagt acatcacaaa cgccgcttac gccgtgaaca acatctccgg
     3241 cctggacaag gatatcatcg gcttcggaaa gttttctacc gtgtacacat acgacgagtt
     3301 caaggatcca gagcaccacc gggccgcttt taacaacaac gacaagctgg gttctaagcg
     3361 caccgccgat ggatccgagt tcgaatcccc taaaaagaaa agaaaggtgg gatcccagct
     3421 gcatttaccg caggttttag ctgacgctgt ctcacgcctg gtcctgggta agtttggtga
     3481 cctgaccgac aacttctcct cccctcacgc tcgcagaaaa gtgctggctg gagtcgtcat
     3541 gacaacaggc acagatgtta aagatgccaa ggtgataagt gtttctacag gaacaaaatg
     3601 tattaatggt gaatacatga gtgatcgtgg ccttgcatta aatgactgcc atgcagaaat
     3661 aatatctcgg agatccttgc tcagatttct ttatacacaa cttgagcttt acttaaataa
     3721 caaagatgat caaaaaagat ccatctttca gaaatcagag cgaggggggt ttaggctgaa
     3781 ggagaatgtc cagtttcatc tgtacatcag cacctctccc tgtggagatg ccagaatctt
     3841 ctcaccacat gagccaatcc tggaagaacc agcagataga cacccaaatc gtaaagcaag
     3901 aggacagcta cggaccaaaa tagagtctgg tcaggggacg attccagtgc gctccaatgc
     3961 gagcatccaa acgtgggacg gggtgctgca aggggagcgg ctgctcacca tgtcctgcag
     4021 tgacaagatt gcacgctgga acgtggtggg catccaggga tcactgctca gcattttcgt
     4081 ggagcccatt tacttctcga gcatcatcct gggcagcctt taccacgggg accacctttc
     4141 cagggccatg taccagcgga tctccaacat agaggacctg ccacctctct acaccctcaa
     4201 caagcctttg ctcagtggca tcagcaatgc agaagcacgg cagccaggga aggcccccaa
     4261 cttcagtgtc aactggacgg taggcgactc cgctattgag gtcatcaacg ccacgactgg
     4321 gaaggatgag ctgggccgcg cgtcccgcct gtgtaagcac gcgttgtact gtcgctggat
     4381 gcgtgtgcac ggcaaggttc cctcccactt actacgctcc aagattacca agcccaacgt
     4441 gtaccatgag tccaagctgg cggcaaagga gtaccaggcc gccaaggcgc gtctgttcac
     4501 agccttcatc aaggcggggc tgggggcctg ggtggagaag cccaccgagc aggaccagtt
     4561 ctcactcacg ggatccaaaa gaactgcgga tggatctgag tttgagagtc caaaaaagaa
     4621 gaggaaggtc ggttctatca acgccatcaa ggctcagtac gacgagttcg ataactttct
     4681 ggataacccc aggctgggct acttcggaca ggctttcttt tctaaggagg gcagaaacta
     4741 catcatcaac tacggaaacg agtgttacga catcctggcc ctgctgagcg gactggccca
     4801 ctgggtggtg gccaacaacg aggaggagtc tcggatcagc cgcacctggc tgtacaacct
     4861 ggacaagaac ctggataacg agtacatctc cacactgaac tacctgtacg acaggatcac
     4921 caacgagctg acaaacagct tctccaagaa ctctgccgct aacgtgaact acatcgctga
     4981 gaccctgggc atcaacccag ctgagttcgc tgagcagtac ttcagatttt ccatcatgaa
     5041 ggagcagaag aacctgggct tcaacatcac aaagctgaga gaagtgatgc tggacagaaa
     5101 ggatatgtcc gagatcagga agaaccacaa ggtgttcgat tctatcagaa ccaaggtgta
     5161 cacaatgatg gactttgtga tctacaggta ctacatcgag gaggatgcca aggtggccgc
     5221 tgccaacaag agcctgcccg acaacgagaa gtctctgagc gagaaggata tcttcgtgat
     5281 caacctgaga ggctccttta acgacgatca gaaggacgct ctgtactacg atgaggccaa
     5341 caggatctgg agaaagctgg agaacatcat gcacaacatc aaggagttcc ggggaaacaa
     5401 gacccgcgag tacaagaaga aggacgctcc aaggctgcct aggatcctgc ctgctggaag
     5461 ggacgtgagc gccttcagca agctgatgta cgccctgaca atgtttctgg acggaaagga
     5521 gatcaacgat ctgctgacca cactgatcaa caagttcgac aacatccagt cttttctgaa
     5581 agtgatgcct ctgatcggcg tgaacgctaa gttcgtggag gagtacgcct tctttaagga
     5641 cagcgccaag atcgctgatg agctgcggct gatcaagtcc tttgccagga tgggagagcc
     5701 aatcgctgac gctaggagag ctatgtacat cgatgccatc cggatcctgg gaaccaacct
     5761 gtcttacgac gagctgaagg ctctggccga caccttcagc ctggatgaga acggcaacaa
     5821 gctgaagaag ggcaagcacg gaatgcgcaa cttcatcatc aacaacgtga tcagcaacaa
     5881 gcggtttcac tacctgatca gatacggcga cccagctcac ctgcacgaga tcgctaagaa
     5941 cgaggccgtg gtgaagttcg tgctgggacg gatcgccgat atccagaaga agcagggcca
     6001 gaacggaaag aaccagatcg accgctacta cgagacctgc atcggcaagg ataagggaaa
     6061 gtccgtgtct gagaaggtgg acgctctgac caagatcatc acaggcatga actacgacca
     6121 gttcgataag aagagatctg tgatcgagga caccggaagg gagaacgccg agagagagaa
     6181 gtttaagaag atcatcagcc tgtacctgac agtgatctac cacatcctga agaacatcgt
     6241 gaacatcaac gctagatacg tgatcggctt ccactgcgtg gagcgcgatg cccagctgta
     6301 caaggagaag ggatacgaca tcaacctgaa gaagctggag gagaagggct ttagctccgt
     6361 gaccaagctg tgcgctggaa tcgacgagac agcccccgac aagaggaagg atgtggagaa
     6421 ggagatggcc gagagagcta aggagagcat cgactccctg gagtctgcta accctaagct
     6481 gtacgccaac tacatcaagt actccgatga gaagaaggcc gaggagttca ccaggcagat
     6541 caacagagag aaggccaaga ccgctctgaa cgcctacctg aggaacacaa agtggaacgt
     6601 gatcatccgg gaggacctgc tgcgcatcga taacaagacc tgtacactgt tcgctaacaa
     6661 ggctgtggcc ctggaggtgg ctcgctacgt gcacgcctac atcaacgaca tcgccgaggt
     6721 gaactcctac tttcagctgt accactacat catgcagagg atcatcatga acgagagata
     6781 cgagaagtct agcggcaagg tgtctgagta cttcgacgcc gtgaacgatg agaagaagta
     6841 caacgataga ctgctgaagc tgctgtgcgt gcctttcgga tactgtatcc cacggtttaa
     6901 gaacctgagc atcgaggccc tgttcgaccg caacgaggct gccaagtttg ataaggagaa
     6961 gaagaaggtg agcggcaact ccggttctaa gcgcaccgcc gatggatccg agttcgaatc
     7021 ccctaaaaag aaaagaaagg tgggatccga ctacaaagac catgacggtg attataaaga
     7081 tcatgatatc gattacaagg atgacgatga caagggatcc aaaagaactg gaagcggagc
     7141 tactaacttc agcctgctga agcaggctgg agacgtggag gagaaccctg gacctgttaa
     7201 catggtgagc aagggcgagg aggataacat ggccatcatc aaggagttca tgcgcttcaa
     7261 ggtgcacatg gagggctccg tgaacggcca cgagttcgag atcgagggcg agggcgaggg
     7321 ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc
     7381 cttcgcctgg gacatcctgt cccctcagtt catgtacggc tccaaggcct acgtgaagca
     7441 ccccgccgac atccccgact acttgaagct gtccttcccc gagggcttca agtgggagcg
     7501 cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc caggactcct ccctgcagga
     7561 cggcgagttc atctacaagg tgaagctgcg cggcaccaac ttcccctccg acggccccgt
     7621 aatgcagaag aagaccatgg gctgggaggc ctcctccgag cggatgtacc ccgaggacgg
     7681 cgccctgaag ggcgagatca agcagaggct gaagctgaag gacggcggcc actacgacgc
     7741 tgaggtcaag accacctaca aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt
     7801 caacatcaag ttggacatca cctcccacaa cgaggactac accatcgtgg aacagtacga
     7861 acgcgccgag ggccgccact ccaccggcgg catggacgag ctgtacaagt aagctcgagc
     7921 ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat
     7981 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     8041 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     8101 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     8161 gggatgcggt gggctctatg gtaccgtcga cctctagcta gagcttggcg taatcatggt
     8221 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     8281 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     8341 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     8401 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     8461 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     8521 tacggttatc cacagaatca ggggataacg caggaaagaa catg
//
